Figure 4: Basic BR_187–385 binds to DNA and forms a saddle-like dimer structure that snuggly fits the acidic double stranded DNA helix.

(A) BR_187–385 concentration-dependent complex formation with a non-specific 270 bp DNA molecule (DNA_270bp) was detected as retarded DNA bands in an electrophoretic mobility shift assay (EMSA). The apparent affinity of the interaction was estimated to around 10 μM. (B) After incubation of BR_187–385 with DNA_270bp, the area of the BR_187–385 monomer peak, obtained at a retention volume of around 17 mL and an absorption wavelength of 280 nm, was slightly reduced. The high-molecular weight population was detected at a retention volume of about 8 mL with an estimated MW of about >1 MDa. DNA_270bp alone would elute at the same retention volume (not shown). (C) The putative BR_187–385-DNA complex was isolated from SEC and run on SDS-PAGE (left) and DNA-Agarose (right) gels. In SDS-PAGE the protein band was detected as a slightly larger complex, possibly retarded by the bound negatively charged DNA. The DNA_270bp that was isolated from SEC was detected at the expected size, and was not retarded due to the low amount of bound BR_187–385. (D) The intermolecular β-sheet dimer resembles a saddle-like structure with an extended intermolecular β-sheet. The electrostatic potentials are plotted in k_bT e_c⁻¹ with the Boltzmann’s constant k_b, the charge of an electron e_c at a temperature T of 298 K. The saddle-like structure snuggly fits an acidic helical non-specific DNA molecule, as predicted in a molecular model of the BR_187–385-DNA complex. (E) Similar to the BR_187–385 dimer, the TATA-box binding protein (TBP, PDB: 1YTB) adopts a saddle-like structure. TBP was superimposed onto the BR_187–387 dimer via shape similarity and represented as white and blue ribbons.